LOCUS       Exported                5938 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Broad host range expression vector pMHE5, complete sequence.
ACCESSION   AY299696
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5938)
  AUTHORS   Fodor BD, Kovacs AT, Csaki R, Hunyadi-Gulyas E, Klement E, Maroti G,
            Meszaros LS, Medzihradszky KF, Rakhely G, Kovacs KL.
  TITLE     Modular broad-host-range expression vectors for single-protein and 
            protein complex purification
  JOURNAL   Appl. Environ. Microbiol. 70 (2), 712-721 (2004)
  PUBMED    14766546
REFERENCE   2  (bases 1 to 5938)
  AUTHORS   Fodor BD.
  TITLE     Direct Submission
  JOURNAL   Submitted (16-MAY-2003) Biophysics, BRC Hungarian Academy of 
            Sciences, Temesvari krt., Szeged, Csongrad 6701, Hungary
REFERENCE   3  (bases 1 to 5938)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..5938
                     /organism="Broad host range expression vector pMHE5"
                     /mol_type="genomic DNA"
                     /db_xref="taxon:244054"
     primer_bind     193..211
                     /label=T7 promoter primer
                     /note="T7 promoter primer"
     promoter        193..211
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      193..209
                     /regulatory_class="promoter"
                     /note="T7 promoter"
     misc_feature    210..235
                     /label=lac operator
                     /note="lac operator"
     protein_bind    212..236
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             251..273
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     regulatory      266..272
                     /regulatory_class="ribosome_binding_site"
     misc_feature    280..282
                     /label=fusion protein start codon
                     /note="fusion protein start codon"
     misc_feature    286..303
                     /label=His-tag
                     /note="His-tag"
     CDS             286..303
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    304..315
                     /label=Factor Xa cleavage site
                     /note="Factor Xa cleavage site"
     CDS             304..315
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     misc_feature    370..393
                     /label=FLAG-tag
                     /note="FLAG-tag"
     CDS             370..393
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    379..393
                     /label=enterokinase cleavage site
                     /note="enterokinase cleavage site"
     misc_feature    397..420
                     /label=Strep-tag II
                     /note="Strep-tag II"
     CDS             397..420
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     primer_bind     complement(474..491)
                     /label=T7 terminator primer
                     /note="T7 terminator primer"
     regulatory      487..534
                     /regulatory_class="terminator"
                     /note="T7 terminator"
     terminator      488..535
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1166..1828
                     /label=pBBR1 oriV
                     /note="pBBR1 oriV"
     CDS             complement(1166..1828)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      1829..2598
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     misc_feature    2822..3815
                     /note="RP4 mob; oriT"
     regulatory      complement(3905..4020)
                     /regulatory_class="terminator"
                     /note="T4 terminator"
     CDS             complement(4280..5071)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside-3:adenyltransferase fusion"
                     /label=aadA
                     /note="streptomycin resistance"
                     /protein_id="AAQ64667.1"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     gene            complement(4280..5071)
                     /gene="aadA"
                     /label=aadA
     regulatory      5807..5922
                     /regulatory_class="terminator"
                     /note="T4 terminator"